Low temperature is a major environmental factor that affects metabolism, growth, development, distribution and production of chilling-sensitive plant, and J. curcas L. is a sustainable energy plant with great potential for biodiesel production due to the fact that its seed contains high oil content, which has attracted much attention worldwide. Our previous work found that the chill hardening improved the chilling tolerance of J. curcas seedlings (Ao et al. in Acta Physiologiae Plantarum 35:153–160, 2013), but its mechanism still remains elusive. In present work, the mechanism of chill hardening-induced chilling tolerance was further investigated in J. curcas seedlings. The results showed that chill hardening at 12 °C for 2 days markedly lowered osmotic and water potentials, which, in turn, maintained relative higher pressure potential in leaves of J. curcas seedlings compared with the control seedlings without chill hardening. In addition, chill hardening gradually increased compatible solutes proline, betaine and total soluble sugar contents compared with the control. When the control and hardened seedlings were subjected to chilling stress at 1 °C for 1–7 days, the chill-hardened seedlings significantly accumulated higher proline, betaine and total soluble sugar contents, which decreased osmotic and water potentials, and maintained higher pressure potential. To further understand the pathways of accumulation of compatible solutes, measurement of activities of ∆1-pyrroline-5-carboxylate synthetase (P5CS), glutamate dehydrogenase (GDH), ornithine aminotransferase (OAT), arginase, proline dehydrogenase (ProDH) and betaine dehydrogenase (BADH) showed that the chill hardening at 12 °C for 2 days obviously increased the activities of P5CS, GDH, OAT, arginase and BADH, as well as lowered ProDH activity both in leaves and stems of J. curcas seedlings to some extent as compared with the control. When the control and hardened seedlings were exposed to chilling stress at 1 °C for 1–7 days, the chill-hardened seedlings generally maintained significantly higher activities of P5CS, GDH, OAT, arginase and BADH. All above-mentioned results illustrated that the chill hardening could induce an accumulation of compatible solutes in leaves of J. curcas seedlings and compatible solutes play important roles in chill hardening-induced chilling tolerance.